I. Field of the Invention
The present invention generally relates to methods and systems for evaluating the length of elongated elements. More particularly, the present invention relates to evaluating the length of elongated elements using, for example, an alternating current stimulus.
II. Background Information
Size-separation and sequencing of chain-like biomolecules (e.g. single stranded deoxyribonucleic acid (DNA), ribonucleic acid (RNA), and proteins) is a process of vital importance in biotechnology and medicine. Sequencing speed is a bottleneck in genomics and allied disciplines. In particular, current DNA sequencing methods involve electrophoretic separation of DNA strands of varying sizes, generated by the well-established polymerase chain reaction (PCR) process. The PCR process generates DNA strands of varying lengths from the original sample, such that the length of a generated strand reflects the identity (e.g. A, C, G or T) of the base at the fluorescently labeled termination position. The sequencing problem is thus reduced to size-separating (or sizing) DNA strands. This step is carried out via electrophoresis in a gel or capillary bundle. The molecules are separated into bands by virtue of difference in their transport rates through the medium as a function of their size, under an applied electric field.
Efforts to substantially raise the throughput rates of these devices are impeded by the “short read length” problem (i.e. inefficient operation at long sequence lengths due to very slow transport of long strands through the medium.) In addition, the use of fluorescence-based optical techniques to detect the DNA bands increases the size and cost of DNA sequencing devices. However, overcoming the limitations of electrophoresis will result in a large technological impact, in terms of new applications such as “personalized medicine” (routine, patient-specific genome sequencing to diagnose genetic health risks), and fast genotyping of new pathogens or biological warfare agents to allow a rapid response.
In recent years, research at the biology-nanotechnology interface has shown potential for creating revolutionary advances in speed, efficiency, reliability, and portability of biomolecule sensors. An underlying advantage of a truly nano-scale biomolecule sensing technique is the ability to detect single molecules at a nanometer length scale and at very short time scales, using only small amounts of sample. Operation at short length and time scales would remove the transport limitations associated with electrophoresis technology. Of several proposed strategies for sizing DNA, the use of nano-scale ion channels is particularly attractive. The sensing element is a nanometer-scale pore (˜2-5 nm in diameter and a few nm long) embedded in a substrate.
Different types of nanopores have been proposed or demonstrated for use in the above devices, and are currently under further development in several research groups. Conventional nanopores have used a direct current (DC) voltage to demonstrate the translocation of biomolecules through the pore. However, more sophisticated sensing protocols are needed to enable the processing of real samples in an efficient and reliable manner, and to optimize, for example, the characteristics of the sensor such as the sensitivity and signal-to-noise ratio.
In view of the foregoing, there is a need for methods and systems for evaluating the length of elongated elements comprising, but not limited to, chain biomolecules mixtures like DNA, RNA, or other proteins, more optimally. Furthermore, there is a need for evaluating the length of elongated elements using, for example, an alternating current stimulus. The elongated elements may comprise, but are not limited to, chain biomolecules mixtures like DNA, RNA, or other proteins.
Moreover, there is a need for methods and systems that include very high sensitivity (single molecule levels), extremely rapid and reversible response due to the short detection length and small time scales (c.a. 1 nm and 1 ms respectively), good signal-to-noise ratio even at low analyte concentrations, as single molecules are detected irrespective of concentration, and concurrent multiple-analyte sensing using arrays of nanopores. These needed methods and systems may increase sizing speeds from 104-105 bases per day on a single electrophoresis instrument, to levels of 107-108 bases per day (e.g. 3-4 orders of magnitude higher.) In addition, there is a need for methods and systems that may be combined with a circuit chip that analyzes the signals from the nanopore array, as well as a micro-fluidic system for handling input and output of analyte samples.